CE-ESI-MS separation of divalent organic and inorganic anions using a tricationic complexing reagent.

A tricationic ion-pairing reagent, 1,3,5-1-butyl-3-methyl-1H-imidazol-3-ium-2,4,6-trimethylbenzene, was used to form complexes with doubly charged anions for their subsequent analysis by capillary electrophoresis-electrospray ionization-mass spectrometry (CE-ESI-MS) in the positive-ion mode. This methodology offers the advantages of greater versatility and sensitivity relative to direct detection of the anions in negative-ion mode, and it can be realized by a number of possible complexation strategies, including pre-column, on-column, and post-column modes. Three model anions, sulfate [SO4²â»], thiosulfate [TSFA, S2O3²â»], and benzenedisulfonate [BZDS, C6H4(SO3)2²â»], were amenable to complexation with the tricationic reagent, yielding singly charged cations with greater mass-to-charge (m/z) ratios than the native analytes. By utilizing optimized parameters obtained through previous work with dicationic reagents and singly charged anions, including the CE separation buffer composition and pH, the concentration of the dicationic reagent, the mode of complexation, the nebulizing gas pressure, and the sheath liquid composition, it was possible to develop a robust CE-ESI-MS method appropriate for the analysis of divalent anions in a mixture.

Association between winter anthocyanin production and drought stress in angiosperm evergreen species.

Leaves of many evergreen angiosperm species turn red under high light during winter due to the production of anthocyanin pigments, while leaves of other species remain green. There is currently no explanation for why some evergreen species exhibit winter reddening while others do not. Conditions associated with low leaf water potentials (Psi) have been shown to induce reddening in many plant species. Because evergreen species differ in susceptibility to water stress during winter, it is hypothesized that species which undergo winter colour change correspond with those that experience/tolerate the most severe daily declines in leaf Psi during winter. Six angiosperm evergreen species which synthesize anthocyanin in leaves under high light during winter and five species which do not were studied. Field Psi, pressure/volume curves, and gas exchange measurements were derived in summer (before leaf colour change had occurred) and winter. Consistent with the hypothesis, red-leafed species as a group had significantly lower midday Psi in winter than green-leafed species, but not during the summer when all the leaves were green. However, some red-leafed species showed midday declines similar to those of green-leafed species, suggesting that low Psi alone may not induce reddening. Pressure-volume curves also provided some evidence of acclimation to more negative water potentials by red-leafed species during winter (e.g. greater osmotic adjustment and cell wall hardening on average). However, much overlap in these physiological parameters was observed as well between red and green-leafed species, and some of the least drought-acclimated species were red-leafed. No difference was observed in transpiration (E) during winter between red and green-leaved species. When data were combined, only three of the six red-leafed species examined appeared physiologically acclimated to prolonged drought stress, compared to one of the five green-leafed species. This suggests that drought stress alone is not sufficient to explain winter reddening in evergreen angiosperms.

CE-ESI-MS analysis of singly charged inorganic and organic anions using a dicationic reagent as a complexing agent.

A dicationic ion-pairing reagent, N,N'-dibutyl 1,1'-pentylenedipyrrolidinium, was used to form complexes with singly charged anions for their subsequent analysis by CE-ESI-MS in positive ion mode. This methodology offers the advantages of greater versatility and sensitivity relative to direct detection of the anions in negative ion mode, and it can be realized by a number of possible complexation strategies, including pre-column, on-column, and post-column modes. Four model anions, perfluorooctanoate, benzenesulfonate (BZSN), monochloroacetate (MCA), and trifluoromethanesulfonimide were amenable to complexation with the dicationic reagent, yielding singly charged cations with greater m/z. By optimizing various parameters, including the CE separation buffer composition and pH, the concentration of the dicationic reagent, the mode of complexation, the nebulizing gas pressure, and the sheath liquid composition, it was possible to develop a robust CE-ESI-MS method appropriate for the analysis of anions in a tap water sample. By this method, LODs were found to be 20.9 and 1.31 ng/mL for MCA and BZSN, respectively.

Enhancement of volatile aglycone recovery facilitated by acid hydrolysis of glucosides from Nicotiana flower species.

Four different Nicotiana flowers (Nicotiana alata (alata), Nicotiana sylvestris (Sy), Nicotiana suaveolens (Su), and Nicotiana tabacum cv. Flue-Cured (FC)) from farms in Virginia and North Carolina were harvested and promptly quenched with liquid nitrogen and hand-ground prior to analysis. Each Nicotiana flower was pre-extracted with hexane to remove unbound volatiles. Fifteen standard compounds that were thought to be in the pre-extract were employed to aid in GC-MS identification and quantification. Glucosides were then chromatographically isolated and next hydrolyzed via 2 M sulfuric acid for 24 h at 75 °C. For each flower, the products of hydrolysis were extracted in tandem with hexane and dichloromethane (DCM) prior to analysis by GC-MS. The mixture of hexane and DCM extracts of the flowers after hydrolysis were then analyzed for each of 15 external standards via GC-MS to determine the concentration of any isolated flower-derived aglycone. Quantitative results for each of the possible 15 free volatile compounds extracted before and after hydrolysis were compared. Benzyl alcohol, phenethyl alcohol, and cis-3-hexenol were found in all Nicotiana both before and after acid hydrolysis. Enormous increases in the mass of benzyl alcohol and phenethyl alcohol were obtained with all flowers as a result of acid hydrolysis. With selected Nicotiana flowers, significant increases were observed for eugenol and cinnamaldehyde. The significant increases observed in cinnamaldehyde and eugenol upon mild acid hydrolysis strongly indicate that this approach could be a viable alternative process for the production scale isolation of these important natural flavor compounds.

Parameters of Reserpine Analogs That Induce MSH2/MSH6-Dependent Cytotoxic Response.

Mismatch repair proteins modulate the cytotoxicity of several chemotherapeutic agents. We have recently proposed a "death conformation" of the MutS homologous proteins that is distinguishable from their "repair conformation." This conformation can be induced by a small molecule, reserpine, leading to DNA-independent cell death. We investigated the parameters for a small reserpine-like molecule that are required to interact with MSH2/MSH6 to induce MSH2/MSH6-dependent cytotoxic response. A multidisciplinary approach involving structural modeling, chemical synthesis, and cell biology analyzed reserpine analogs and modifications. We demonstrate that the parameters controlling the induction of MSH2/MSH6-dependent cytotoxicity for reserpine-analogous molecules reside in the specific requirements for methoxy groups, the size of the molecule, and the orientation of molecules within the protein-binding pocket. Reserpine analog rescinnamine showed improved MSH2-dependent cytotoxicity. These results have important implications for the identification of compounds that require functional MMR proteins to exhibit their full cytotoxicity, which will avoid resistance in MMR-deficient cells.